The term “Nuclease” means an all-inclusive term of nucleolytic enzymes that specifically degrades nucleic acids. When a nuclease reacts with nucleic acids such as deoxyribonucleic acid and ribonucleic acid, a phosphodiester bond between sugar and phosphate in the nucleic acids is hydrolyzed, and nucleosides are generated.
Nucleases are categorized as one type of ester hydrolase whose EC number (Enzyme Commission number) is EC. 3.1. Moreover, the nucleases are categorized into ribonucleases, which decompose RNA, and deoxyribonucleases, which decompose DNA. Furthermore, the nucleases may be categorized according to the type of decomposition.
An enzyme that serves as a catalyst to cleavage nucleic acids from the inside (endo-) of a sequence of the nucleic acids is called endonuclease. Various restriction enzymes are among typical endonucleases. While, an enzyme that serves as a catalyst to cleavage nucleic acids in such a way as to shave from a 5′ or 3′ end of the nucleic acids from the outside (exo-) to the inside of a sequence of the nucleic acids is called exonuclease. Exonuclease III and the like are known as typical exonucleases.
Nucleases are now used in various scenes from laboratory scale to industrial scale. For example, the use of nucleases makes it possible to decrease the viscosity of cell extracts because of nucleic-acid degradation activity thereof. Accordingly, if nucleases are used when proteins and other objective substances in the cell extracts are isolated and purified, the following advantages can be expected: shortening of process time, an improvement in the amount of objective substances obtained, an improvement in fractionation by centrifugal separation method (isolation of pellet and supernatant), smooth filtration of a solution (particularly ultrafiltration), an improvement in the efficiency of chromatographic process and the like. Moreover, if nucleases are used when viruses, inclusion bodies, or the like to which nucleic acids are nonspecifically adsorbed are isolated and purified, an improvement can be expected to be made in the yield of the above entities. Furthermore, if nucleases are used in a process of preparing samples used in assays such as ELISA, chromatography, 2D-PAGE, and footprint analysis for the analysis of biological samples, it is possible to avoid measurement error associated with unnecessary nucleic acids.
As a nuclease that can be used in such various scales or scenes, an endonuclease derived from Serratia spp (Serratia spp.) is known (See the specification of Japanese Patent No. 2604365 and the specification of U.S. Pat. No. 5,173,418 as Patent Documents 1 and 2, respectively; the contents of Patent Documents 1 and 2 are incorporated herein by reference). Microorganisms of the genus Serratia may contain disease-causing bacteria as opportunistic infection bacteria. Therefore, the endonuclease disclosed in Patent Document 1 is produced as an extracellular secretion-type enzyme, which is secreted outside a cell, with the use of Escherichia coli by gene-recombination technology. Incidentally, the endonuclease derived from Serratia spp disclosed in Patent Document 1 is marketed under the brand of Benzonase (Registered Trademark) (See, as Non-Patent Document 1, “Benzonase—a unique endonuclease—,” [online], Jan. 1, 2008, Merck Ltd., [Searched on Jul. 30, 2010], Internet <URL: http://www2.merck.co.jp/japan/chemical/pdf/info_pdf/071225_Ben zonase—16p.pdf>”; the contents of Non-Patent Document 1 are incorporated herein by reference).
Example of nucleases derived from non-pathogenic microorganisms, a nuclease produced by bacteria of the genus Streptomyces (Streptomyces spp.), which is one type of actinomycetes, is known (See the following documents as Non-Patent Documents 2 to 7: Biochem. J. 1995 306, 93-100; Biochem. J. 1992 281, 231-237; Appl Microbiol Biotechnol. 1995 November; 43(6): 1056-1060; Biochimica et Biophysica Acta (BBA)—General Subjects, Volume 1721, Issues 1-3, 18 Jan. 2005, 116-123; FEMS Microbiology Letters, Volume 237, Issue 2, 15 Aug. 2004, 273-278; and Process Biochemistry Volume 40, Issues 3-4, March 2005, 1271-1278; the contents of Non-Patent Documents 2 to 7 are incorporated herein by reference). The nucleases disclosed in Non-Patent Documents 2 and 3 are produced as intracellular accumulation-type enzymes. Whereas, the nucleases disclosed in Non-Patent Documents 4 to 7 are extracellular secretion-type enzymes which are secreted out of cells.